Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Gonadotropin secretion and gene expression are differentially regulated by hypothalamic GnRH pulses by unknown mechanisms. GnRH stimulates calcium influx through L-type voltage-gated channels and activates phospholipase C, leading to increased protein kinase C (PKC) and mitogen-activated protein kinase activity. We found differential contributions of these pathways to GnRH-stimulated rat LH subunit transcription in pituitary gonadotropes and cell lines. Endogenous transcription of the alpha- and LHbeta-subunits in rat pituitary cells was stimulated by GnRH. Independent PKC activation by phorbol myristate acid stimulated only the alpha-subunit gene. In contrast, an L-channel antagonist (nimodipine) inhibited only LHbeta stimulation by GnRH, and an L-channel agonist (BayK 8644) stimulated only basal LHbeta transcription. GnRH induction of a rat alpha-subunit promoter construct in alphaT3 cells was unaffected by nimodipine or elimination of external calcium, while both treatments eliminated the LHbeta response. Application of a mitogen-activated kinase kinase (MEK) inhibitor (PD098059) decreased basal and GnRH-stimulated alpha-subunit promoter activity and had no effect on LHbeta promoter activity. In pituitary cells from mice bearing an LHbeta promoter-luciferase reporter transgene, GnRH stimulation was inhibited by nimodipine but not by PD098059. Thus, GnRH induction and basal control of the alpha-subunit gene seem to occur through the PKC/mitogen-activated protein kinase pathway, while induction of the LHbeta gene is dependent on calcium influx. Differential signaling from the same receptor may be a mechanism for preferential regulation of transcription.
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PMID:Differential gonadotropin-releasing hormone stimulation of rat luteinizing hormone subunit gene transcription by calcium influx and mitogen-activated protein kinase-signaling pathways. 951 61

The regulation of LH and FSH subunit gene expression is under the control of GnRH. Physiological changes in the frequency of pulsatile GnRH release from the hypothalamus result in differential stimulation of alpha-, LHbeta-, and FSHbeta-gene expression. Previous studies indicate that the GnRH receptor couples to G proteins of the G(q/11) family, with phosphoinositide turnover and its resultant increase in intracellular calcium concentration and protein kinase C (PKC) activation, to stimulate secretion of LH and FSH. However, the molecular mechanisms by which GnRH mediates its transcriptional effects remain largely unknown. We used GH3 cells, constitutively expressing the rat GnRH receptor (GGH(3)-1' cells) and transiently transfected with a luciferase reporter gene controlled by either the alpha, LHbeta, or FSHbeta gene regulatory region (alphaLUC, LHbetaLUC, and FSHbetaLUC, respectively), to examine the roles of several signal transduction pathways in the GnRH-mediated stimulation of gonadotropin subunit gene expression. Activation of PKC by phorbol, 12-myristate, 13-acetate resulted in an increase in the luciferase activity of all three gonadotropin subunit gene reporter constructs. Phorbol, 12-myristate, 13-acetate had a greater stimulatory effect, relative to the maximal stimulation with GnRH, for the beta-subunit genes than for the alpha-subunit gene. Depletion of PKC, or inhibition of PKC by GF109203X, demonstrated that PKC-dependent pathways play a larger role in the GnRH-mediated transcriptional control of the LHbeta- and FSHbeta-genes than the alpha-subunit gene. In contrast, an L-type calcium channel agonist, Bay K 8644, was able to stimulate alphaLUC but not LHbetaLUC or FSHbetaLUC. Nimodipine, an L-type calcium channel antagonist, had a larger inhibitory effect on the GnRH response of alphaLUC, relative to LHbetaLUC or FSHbetaLUC. We conclude from these results that the differential regulation of gonadotropin subunit gene expression by GnRH is caused, in part, by differential use of signal transduction pathways, activated upon GnRH binding.
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PMID:Differential use of signal transduction pathways in the gonadotropin-releasing hormone-mediated regulation of gonadotropin subunit gene expression. 952 69

Expression of the LHbeta gene has been shown to be modulated by both the orphan nuclear receptor, steroidogenic factor-1 (SF-1), and the early growth response protein 1, Egr-1. It is also well known that LHbeta mRNA levels are increased after hormonal activation of the protein kinase C (PKC) signaling system, for example by GnRH; however, the mechanisms by which the PKC system exerts this effect has not been fully characterized. By transient transfection of the GH3 cell line, we demonstrate that activation of the PKC system with the phorbol ester, phorbol 12-myristate 13-acetate (PMA), increases activity of region -207/+5 of the rat LHbeta gene promoter (approximately 2-fold) and markedly augments SF-1-induced stimulation (95-fold in the presence of both factors vs. 13-fold for SF-1 alone). Mutation of the two previously identified Egr-1 sites not only prevents Egr-1 effects on the LHbeta gene promoter, but also eliminates the synergistic response to PMA and SF-1 together, findings that were confirmed in a longer construct spanning region -797/+5. In the gonadotrope-derived cell line, alphaT3-1, these mutations eliminate the GnRH responsiveness of the -207/+5 LHbeta promoter construct. We next show that PMA treatment (GH3 and alphaT3-1 cells) or GnRH treatment (alphaT3-1 cells) induces expression of Egr-1, as detected by Egr-1 interaction with Egr-1 DNA-binding sites in the rat LHbeta gene promoter sequence. Furthermore, we demonstrate that PMA increases steady-state Egr-1 mRNA levels via increased Egr-1 transcription. We conclude that PMA-induced stimulation of LHbeta gene expression is achieved, at least in part, by induction of Egr-1 expression.
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PMID:The protein kinase C system acts through the early growth response protein 1 to increase LHbeta gene expression in synergy with steroidogenic factor-1. 989 16

GnRH regulation of LH secretion is well understood and involves Ca(2+) mobilization. However, the mechanism by which GnRH activates transcription of the LHbeta gene is controversial. GnRH is known to elevate intracellular calcium and activate the protein kinase C (PKC) pathway. The present study evaluated the pathway(s) involved in GnRH induction of LHbeta transcription. We have previously reported that the equine LHbeta (eLHbeta -448/+60) promoter is active in alphaT3-1 cells. Therefore, we created a clonal, stably transfected alphaT3-1 gonadotroph cell line harboring the eLHbeta promoter (-448/+60) fused to the luciferase reporter gene. Administration of a GnRH agonist resulted in induction of promoter activity that was completely inhibited by the antagonist antide. Various calcium-affecting drugs had no effect on the promoter. Administration of phorbol 12-myristate 13-acetate (PMA) elicited an activation similar to, albeit lower than, that with GnRH. Down-regulation or pharmacological inhibition of PKC completely blocked PMA's induction of the promoter, while GnRH induction was only partly attenuated. Treatment with the mitogen-activated protein kinase (MAPK) kinase inhibitor, PD98059, completely inhibited the activation of eLHbeta by PMA but only partly diminished GnRH's induction. Expression of the transcription factor, early growth response protein 1 (Egr1), correlated completely with activation of MAPK, suggesting that Egr1 is the factor through which PKC/MAPK acts. Our data suggest that GnRH induces activity of the eLHbeta promoter by activating a signal transduction cascade involving PKC-MAPK-Egr1 but that has no significant requirement for calcium.
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PMID:Gonadotropin-releasing hormone activates the equine luteinizing hormone beta promoter through a protein kinase C/mitogen-activated protein kinase pathway. 1045 49

A steroidogenic tilapia gonadotropin (taGtH=LH) was purified from pituitaries of hybrid tilapia (Oreochromis niloticus x O. aureus) and a homologous RIA was established. This RIA enabled the study of the endocrine regulation of GtH release, the transduction pathways involved in its secretion and its profile during the spawning cycle. Discrepancies between steroid and taGtH peaks during the cycle led to the conclusion that an additional gonadotropin similar to salmonid FSH operates early in the cycle. In order to identify this hormone and to study the endocrine control of synthesis of all gonadotropin (GtH) subunits, a molecular approach was taken. The cDNA sequences and the entire gene sequences encoding the FSHbeta and LHbeta subunits, as well as an incomplete sequence of the glycoprotein hormone alpha subunit (GPalpha), were cloned. Salmon gonadotropin-releasing hormone (sGnRH) elevated mRNA steady-state levels of all three GtH subunits in cultured pituitary cells. Pituitary adenylate cyclase-activating polypeptide (PACAP) and neuropeptide Y (NPY) also stimulated the expression of these subunits and potentiated the effect of GnRH, except that NPY did not affect FSHbeta. The GnRH and NPY effects were found to be mediated mainly through protein kinase C (PKC), while protein kinase A (PKA) cascade was involved to a lesser extent. Mitogen-activated protein kinase (MAPK) cascade takes part in mediating GnRH effects, possibly via PKC. Testosterone (T) and estradiol (E2), but not 11-ketotestosterone (KT), are able to elevate GPalpha and LHbeta mRNAs in pituitary cells of early maturing or regressing males. Low levels of T exposure are associated with elevated FSHbeta mRNA in cells of mature fish, while higher levels suppress it, but elevate LHbeta mRNA. In vivo observations also showed the association of low T levels with increased FSHbeta mRNA and high T levels with elevated LHbeta mRNA. In accordance with these findings, analysis of LHbeta and FSHbeta 5' gene-flanking regions revealed on both gene promoters a GtH-specific element (GSE), half site estrogen response elements (ERE), cAMP response element (CRE) and AP1. In vitro experiments showed that recombinant human activin-A leads to higher levels of GPalpha, FSHbeta and LHbeta mRNAs in pituitary cell culture. Porcine inhibin marginally decreased the mRNA levels of GPalpha and FSHbeta, but at a low level (1 ng/ml) it stimulated that of LHbeta. These results shed some light on certain hypothalamic and gonadal hormones regulating the expression of GtH subunit genes in tilapia. In addition, they provide evidence for their differential regulation, and insight into their mode of action.
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PMID:Regulation of gonadotropin subunit genes in tilapia. 1139 84

The role of mitogen-activated protein kinase (MAPK, also known as extracellular signal regulated kinase; ERK) stimulation in gonadotropin-releasing hormone (GnRH) signaling was investigated in cultured pituitary cells of tilapia hybrids (Oreochromis niloticus x O. aureus). Exposure of the cells to salmon GnRH (sGnRH) resulted in a dose- and time-dependent elevation in ERK levels. The PKC activator, 1-O-tetradecanoyl phorbol-13-acetate (TPA) increased kinase levels, while addition of GnRH had no further effect. However, chronic exposure to TPA resulted in reduction of basal and GnRH-induced ERK elevation. When PKC was inhibited by GF109203X, the GnRH-elevated ERK levels were totally abolished. The role of MAPK activation on GPalpha, FSHbeta and LHbeta gene expression was determined by administration of MAPK-kinase (MEK) inhibitor (PD98059; PD). This inhibitor completely blocked GnRH-induced increases in ERK activity. Furthermore, it suppressed GPalpha and LHbeta mRNA responses to GnRH, but had no effect on FSHbeta transcript levels. PD also decreased basal LHbeta mRNA levels. These results indicate that in tilapia pituitary cells, GnRH activates MAPK cascade in a PKC-dependent manner. ERK is involved in GnRH elevation of GPalpha and LHbeta, but not in FSHbeta genes transcription.
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PMID:GnRH receptor signaling in tilapia pituitary cells: role of mitogen-activated protein kinase (MAPK). 1139 87

Gonadotropin-releasing hormone (GnRH) stimulates gonadotropin (GTH) subunit gene expression via G protein-coupled membrane receptors. GnRH-stimulated GTH subunit gene expression is mediated by protein kinase C (PKC) and Ca(2+) signaling pathways. Recent numerous reports on signal transduction pathways which are involved in GnRH stimulation of mammalian GTH subunit genes showed differential sensitivity of GTH subunit genes to the two signaling pathways. Our recent studies on salmon GTH (sGTH) IIbeta subunit gene showed that its stimulation by GnRH is dependent on the PKC pathway. Furthermore, gel retardation and mutagenesis studies suggested that pituitary homeo box 1 (Ptx1) and Sp1 mediate the GnRH-induced PKC signaling on the sGTHIIbeta gene. However, both PKC and Ca(2+) pathways are involved in the GnRH-stimulated GTH alpha and LHbeta genes. Different preference to the pathways were often reported in a certain GTH subunit gene in different circumstances, suggesting that molecular targets of the two signaling pathways are different. Ets-related factor and cAMP response element binding protein have been proposed as targets of GnRH signaling on GTH alpha genes. Sp1 and early growth response protein 1 play pivotal roles in GnRH-stimulated LHbeta gene expression in synergism with steroidogenic factor-1 and Ptx1. Activating protein-1 mediates GnRH-induced PKC signaling to stimulate FSHbeta gene expression. Therefore, divergent transcription factors are involved in GnRH stimulation of GTH subunit gene expression, and molecular mechanisms of GnRH stimulation may be partially conserved between sGTH IIbeta and mammalian LHbeta genes.
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PMID:Signal transduction pathways and transcription factors involved in the gonadotropin-releasing hormone-stimulated gonadotropin subunit gene expression. 1139 88

Detailed studies have been focused on the mechanisms by which the rat alpha and LHbeta genes are differentially regulated by GnRH and indicate that differential sensitivity to the second messenger exists in a physiological context. Differential signaling from the GnRH receptor may be a mechanism for preferential regulation of luteinizing hormone subunit gene transcription; however which of these genes are specifically regulated by PKC or calcium and how GnRH pulsatility could preferentially activate individual pathways of second messengers within gonadotrope cells remain unclear. Several transcription factors that have profound effects on basal and/or GnRH-stimulated LHbeta gene promoter activity have been identified: SF-1, Egr-1, Sp-1. A model explaining possible interactions among them in mediating GnRH responsiveness of the LHbeta gene has been proposed: Sp1, SF-1 and Egr-1 form a tripartite GnRH response element which is sensitive to the spacing changes between the upstream Sp1 binding sites and the downstream SF-1/Egr-1 binding elements and SF-1 plays a critical role in integrating the effects of Sp1 and Egr-1. GnRH responsive element located on LHbeta gene promoter in position between -495 to -342 has been identified. At 3'-end of the promoter three Sp-1 binding sites have been identified: position -416, sequence: GGGGGCTGGG and two sites almost completely overlapping, position -403, sequence; GGGGCGGCGCCCA while at the 5'region of the promoter one Sp-1 binding site exists: position -450, sequence: ACCACACCCATTTTTGG. The 5'Sp1 site overlaps a CArG box (at -443 to -434, sequence: CCATTTTTGG) which seems to be essential in LHbeta gene sensitivity for pulsatile GnRH stimulation.
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PMID:GnRH pulsality and the differential activation of the rat luteinizing hormone subunit genes in the anterior pituitary gland. 1178 41

Pulsatile secretion of GnRH is the major regulator of gonadotropin (LH, FSH) gene expression and secretion. Recently, GnRH has been shown to rapidly stimulate the expression of early growth response protein-1 (Egr-1), a transcription factor that is essential for LHbeta gene expression in the pituitary. In this study, we examined the regulatory elements and signal transduction pathways by which GnRH regulates Egr-1 transcription. Deletion analysis of the murine Egr-1 promoter identified two regions (-370 to -342 and -116 to -73) that are critical for GnRH responsiveness in alphaT3 pituitary gonadotrope cells. The first region, which contains two serum response elements (SREs), contributed about 70-80% of GnRH inducibility, whereas the second region, which contains two SREs and one Ets binding site, conferred an additional 20-30% of activity. Mutations that abolish protein binding to these SREs and Ets binding sites completely eliminated GnRH-mediated transcriptional activation of the Egr-1 promoter. Mutation of cAMP response element reduced promoter activity by 40%. Using specific protein kinase inhibitors, GnRH stimulation of Egr-1 expression was found to be dependent on PKC/ERK pathways. In addition, GnRH activated p90 ribosomal S6 kinase, which has the potential to phosphorylate serum response factor and cAMP response element binding protein. We conclude that GnRH stimulation of Egr-1 gene expression requires several distinct SREs/Ets elements and a cAMP response element and is mediated via activation of PKC/ERK signaling pathways.
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PMID:GnRH regulates early growth response protein 1 transcription through multiple promoter elements. 1181 96

The role of ERK and Jun N-terminal kinase (JNK) in basal- and GnRH-stimulated LHbeta-promoter activity was examined in the gonadotroph cell line LbetaT-2. GnRH agonist (GnRH-A) stimulates the MAPK cascades ERK, JNK, and p38MAPK, with a peak at 7 min for ERK and at 60 min for JNK and p38MAPK. The rat glycoprotein hormone LHbeta-subunit promoter, linked to the chloramphenicol acetyl transferase (CAT) reporter gene, was used to follow its activation. Addition of GnRH-A (10 nM) to LbetaT-2 cells resulted in a 6-fold increase in LHbeta-CAT activity at 8 h, which was markedly reduced by a GnRH antagonist. The PKC activator 12-O-tetradecanoylphorbol-13-acetate (TPA), but not the Ca(2+) ionophore ionomycin, stimulated LHbeta-CAT activity. Addition of GnRH-A and TPA together did not produce an additive response. Down-regulation of PKC, but not removal of Ca(2+), abolished the GnRH-A and the TPA response. Cotransfection of the LHbeta-promoter and the constitutively active form of Raf-1 stimulated basal and GnRH-A-induced LHbeta-CAT activity. The dominant negative forms of the ERK cascade members Ras, Raf-1, and MAPK/ERK kinase (MEK) markedly reduced basal and GnRH-A-induced LHbeta-CAT activity, Similar results were obtained with the MEK inhibitor PD 098059. Cotransfection of the LHbeta-promoter and the constitutively active CDC42 stimulated basal and GnRH-A-induced LHbeta-CAT activity. The dominant negative forms of the JNK cascade members Rac, CDC42, and SEK markedly diminished basal and GnRH-A-induced LHbeta-CAT activity. Interestingly, the constitutively active form of c-Src stimulated the basal and the GnRH-A response, whereas the dominant negative form of c-Src, or the c-Src inhibitor PP1 diminished basal and the GnRH-A response. We conclude that ERK and JNK are involved in basal and GnRH-A stimulation of LHbeta-CAT activity. c-Src participates also in LHbeta-promoter activation by a mechanism which might be linked to ERK and JNK activation.
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PMID:Activation of MAPK cascades by GnRH: ERK and Jun N-terminal kinase are involved in basal and GnRH-stimulated activity of the glycoprotein hormone LHbeta-subunit promoter. 1186 27


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